Abstract
Successful delivery of plasmid DNA into the microbial cells is fundamental in recombinant DNA technology. Natural bacterial transformation is limited to only certain species due in part to the repulsive forces between negatively charged DNA and bacterial membranes. Most common method of DNA delivery into bacteria is artificial transformation through heat shock and electroporation. These methods require sophisticated instruments and tedious steps in preparation of competent cells. Transformation by conjugation is also not applicable to all plasmids. Nanoparticles have been used successfully in therapeutics for drug delivery into animal cells. They are starting to gain popularity in plant sciences as novel DNA nano carriers. Despite their promise as tool for DNA delivery, their use in microbial cell transformation has not been reported yet. Here we report the synthesis of carbon dots (CDs) from citric acid and β-alanine and their use in DNA delivery into E. coli cells. CDs were fabricated using microwave assisted synthesis. Plasmids carrying RFP reporter and ampicillin resistance genes were transferred to bacterial cells and further confirmed using polymerase chain reaction. Our findings indicate that CDs can be used successfully for delivery of foreign DNA of up to 10 kb into E. coli. We have demonstrated the use of β-alanine/citric acid carbon dots as nanocarriers of DNA into E. coli cells and identified their limitation in terms of the size of plasmid DNA they could carry. Use of these carbon dots is a novel method in foreign DNA delivery into bacterial cells and have a potential for the transformation of resistant organism for which there is still no reliable DNA delivery systems.
Highlights
Successful delivery of plasmid DNA into the microbial cells is fundamental in recombinant DNA technology
We report the effect of light and temperature on carbon dots potential for plasmid DNA delivery into E. coli cells
Our results indicate higher number of transformed colonies in cells incubated at 4 °C followed by 25 °C and 37 °C (Fig. 6) which could be due to the higher activity of E. coli cells at higher temperature (37 °C) that leads to higher release of reactive oxygen species (ROS) and cell toxicity
Summary
Successful delivery of plasmid DNA into the microbial cells is fundamental in recombinant DNA technology. We have demonstrated the use of β-alanine/citric acid carbon dots as nanocarriers of DNA into E. coli cells and identified their limitation in terms of the size of plasmid DNA they could carry. Use of these carbon dots is a novel method in foreign DNA delivery into bacterial cells and have a potential for the transformation of resistant organism for which there is still no reliable DNA delivery systems. Carbon dots with high chemical stability, auto-fluorescence, customizable surfaces, ease of synthesis and minimal negative impacts on the environment are gaining popularity for gene delivery in humans, animals and even plants[5,6,8,14,15] Their use as nanocarriers in microbials transformation has not been reported yet. We report the effect of light and temperature on carbon dots potential for plasmid DNA delivery into E. coli cells
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